Validation and Implementation of Methods inAssessing Job-Related Exposures in Dairy Farm Workers

Lorena Ray, Christina Rohlik, Jung-Keun Lee, Nicky Beck, Scott MeschkeDepartment of Environmental and Occupational Health Sciences

• Salmonella and E. coli were identified on the Dairy Farm.

• Although organisms grew using the other methods of detection,Salmonella and E. coli were not detected using the SKC Biosampler.Consequently, it appears that the SKC Andersen is a more appropriatemethod for detecting these organisms in an agricultural setting.

Cattle are sources of bacteria; as a result, dairy farm workers are ata greater risk of exposure to infectious bacteria such asCampylobacter, E. coli O157:H7, and Salmonella. Yakima County isthe leading dairy producing region within Washington State and hasthe largest inventory of cattle in any county in Washington. YakimaCounty has case rates for Campylobacter and Salmonella as high as2 to 3 times the state of Washington and the case rate of E. coliO157:H7 has periodically exceeded Washington’s rates. The primarygoal of this research project was to determine the environmentallevels of these potential pathogens in the dairy setting to whichworkers may be exposed. Several routes of exposure on the DairyFarm were considered significant including air surfacecontamination. Possible air borne bacterial pathogens were detectedusing the SKC Andersen 6-stage, the SKC Andersen single stage,and the SKC Biosampler. Swabs and contact plates with selectiveand non-selective media were used to sample different surfaces onthe farm. Organisms grew on the swabs and both E. coli andSalmonella were detected within in the SKC Andersen six-stage,SKC Andersen single stage, and contact plates. These organisms,however, have not been identified using the SKC Biosampler.

Introduction• Despite relatively little information on the job-related routes ofexposure to bacterial pathogens, there is considerable evidencedemonstrating enteric illness resulting from direct or indirect farmanimal contact.

• Enteric illness can cause severe diarrhea.• The frequency in outbreaks of zoonotic diseases has beenincreasing, and can result in hospital visitations, missed days of work,loss of productivity, and financial burden.• Zoonotic bacteria like Campylobacter spp., Salmonella spp., and E.coli O157:H7 are common causative agents leading to enteric illnessin humans.

• There are numerous potential routes for exposure for farm workerson cattle farm to Campylobacter spp., Salmonella spp., and E. coli,including contaminated clothing or surfaces, vehicles, water,aerosols, fomites and direct contact with animals or animal wastes.[1]• Campylobacter spp. has been recently described to be widelydistributed on cattle farms in the State of Washington with aprevalence of 23% to 47%. [2]

• Yakima County is primarily an agricultural area, with approximatelyhalf devoted to fruits and vegetables and the other half devoted tocattle and dairy farms.

• Yakima County is also the leading dairy producing region within thestate and has the largest inventory of cattle and sheep in any countyin Washington.

• As a result of the labor intensive agriculture in the county, there arelarge populations of minorities, primarily Hispanic, and migrant farmworkers.

• Yakima County has a history of consistently elevated case rates forCampylobacter and Salmonella, 2 to 3 times Washington’s rates. [3]• The case rate of E. coli O157:H7 in Yakima County has periodicallyexceeded the State case rates. [4]

Materials and Methods

Field Study Sampling:

Bioaerosols-

• The SKC Biosampler was run for 30minutes in two different barns.

• Bacterial pathogens were detectedusing the SKC Andersen 6-stage andthe SKC Andersen single stage. Thestages were run for 30 seconds, 45seconds, 1 minute, 1.5 minutes, and 2minutes.

Surfaces-

• Swabs and contact plateswith selective and non-selectivemedia were used to sampledifferent surfaces (barn gates,clothing, car tires, tractorhandles, and water troughs) onthe farm.

• The contact plates werepressed firmly on each surfacefor 30 seconds. Swabs werepre-moistened with Tween 80and stored in PBS aftersampling.Laboratory Study:

Contact plates and SKCAndersen stages-

• The samples were incubated for24 hours and then wereprocessed in the laboratory forbacterial culture on selective andnon-selective media.

• The resulting bacterial growthwas enumerated, colony purified,morphologically characterized,and its identity confirmed by anAPI strip.

Swabs and SKC Biosampler-

• The samples were vortexed and then filtered. The filter plates werethen incubated for 24 hours.

• The resulting bacterial growth was enumerated, the colony purified,its morphology characterized, and its identity confirmed by an API strip.

Media Used:▪ Difco Nutrient Agar ▪ Remel Brilliant Green Agar Modified▪ Difco MI Agar ▪ Difco Actinomycete Isolation Agar▪ Difco MacConkey Agar

Fig. 2 SKC Andersen 6-stage

Fig. 3 Filter Unit

Fig. 1 SKC Biosampler

• E. coli was isolated and confirmed using the SKC Andersen 6-stage, the SKC Andersen single stage, the swabs, and the contactplates.

• E. coli was confirmed at 99.6% using the API strips.

• Salmonella spp. was detected but not confirmed using the SKCAndersen 6-stage, the SKC Andersen single stage, and the contactplates.

• We have not identified Salmonella, Campylobacter, and E. coliusing the SKC Biosampler.

Results/Conclusions

Future Study• Assessing the para-occupational (take-home) exposure pathwaysfor Campylobacter spp., Salmonella spp., and E. coli.

• Archiving the samples at -80°C and looking for antimicrobialresistant genes.

AcknowledgementsSpecial thanks to: The University of Washington Department of Environmental and OccupationalHealth Sciences, the Environmental and Occupational Health Microbiology Lab, Yakima County, theNational Institute of Environmental Health Sciences (NIEHS) grant number: 1R25ES016150 - 01.This research was made possible by funding through the Pacific Northwest Agricultural Safety andHealth Center, CDC/NIOSH #2 U50 OH07544.

References[1] Cole, D. J., V. R. Hill, F. J. Humenik, and M.D. Sobsey. 1999. Health, safety, andenvironmental concerns of farm animal waste. Occup Med 14:423-48.

[2] Smith, K. E., S. A. Stenzel, J. Bender, E. Wagstrom, D. Soderlund, F. T. Leano, C. M. Taylor,P. A. Bellelsle, and R. Danila. 2004. Outbreaks of enteric infections caused by multiple pathogensassociated with calves at a farm day camp. Pediatr Infect Dis J 23:1098-104.

[3] Baer, Rebecca, et al. 2004. Washington State Communicable Disease Report. WashingtonState Department of Health 20-62.

[4] Proctor, M. E., and J. P. Davis. 2000. Escherichia coli O157:H7 infections in Wisconsin, 1992-1999. Wmj 99:32-7.

Brilliant

Green Agar

Nutrient

Agar

MacConkey

AgarMI Agar AIA Agar

6 - Stage 52 4638 152 * ND

Single Stage 18 7360 302 ND 7720

Contact Plate ND 3327 544 * 3816

Swabs * 6453 1154 * 116

Biosampler * 598 ND ND 3

Table 1: Average Number of Colonies Detected for Four Sampling Trips

Table 2: Agent Containing Particles Per Unit of Area Sampled

Abstract

Description of the Field Site

• = This agar was not used ND = Not Detected

E. coli

Total

Coliforms6 - Stage

(ACPLA) 0.59 27.04 1.03 * * 0.51

Single Stage

(ACPLA) 0.37 24.72 1.03 ND ND 0.55

Contact Plate

(Avg. CFU /

in^2) ND 60.08 6.04 * * 90.07

Swabs (Avg.

CFU/cm2) * * * 8.99 6.57 *

Biosampler

(Avg. CFU / L) * 0.27 ND * * ND

MacConkey

Agar

MI AgarAIA Agar

Brilliant

Green Agar

Nutrient

Agar